Citrinin-Generated Reactive Oxygen Species Cause Cell Cycle Arrest Leading to Apoptosis via the Intrinsic Mitochondrial Pathway in Mouse Skin

Abstract

The mycotoxin, citrinin (CTN), is a contaminant of various food and feed materials. Several in vivo and in vitro studies have demonstrated that CTN has broad toxicity spectra; however, dermal toxicity is not known. In the present investigation, dermal exposure to CTN was undertaken to study oxidative stress, DNA damage, cell cycle arrest, and apoptosis in mouse skin. A single topical application of CTN caused significant change in oxidative stress markers, such as lipid peroxidation, protein carbonyl content, glutathione (GSH) content, and antioxidant enzymes in a dose-dependent (25–100 μg/mouse) and time-dependent (12–72 h) manner. Single topical application of CTN (50 μg/mouse) for 12–72 h caused significant enhancement in (1) reactive oxygen species (ROS); (2) cell cycle arrest at the G0/G1 phase (30–71%) and G2/M phase (56–65%) along with the induction of apoptosis (3.6–27%); (3) expression of p53, p21/waf1; (4) Bax/Bcl₂ ratio and cytochome c release; and (5) activities of caspase 9 (22–46%) and 3 (42–54%) as well as increased poly(ADP-ribose) polymerase cleavage. It was also observed that pretreatment with bio-antioxidants viz butylated hydroxyanisole (55 μmol/100 μl), quercetin (10 μmol/100 μl), or α-tocopherol (40 μmol/100 μl) resulted in decreases of ROS generation, arrest in the G0/G1 phase of the cell cycle, and apoptosis. These data confirm the involvement of ROS in apoptosis and suggest that these bio-antioxidants may be useful in the prevention of CTN-induced dermal toxicity.

citrinin, mouse skin, ROS, apoptosis, antioxidants